Ex vivo culture of chimeric antigen receptor T cells generates functional CD8+ T cells with effector and central memory-like phenotype

The anti-tumor efficacy of adoptively transferred T cells requires their in vivo persistence and memory polarization. It is unknown if human chimeric antigen receptor (CAR)-expressing T cells can also undergo memory polarization. We examined the functional status of CAR CD8(+) T cells, re-directed to Lewis Y antigen (LeY-T), throughout a period of ex vivo expansion. Immediately before culture CD8(+) T cells comprised a mixture of phenotypes including naive (CD45RA(+)/CCR7(+)/CD27(+)/CD28(+)/perforin-), central memory (CM, CD45RA /CCR7(lo)/CD27(+)/CD28(+)/perforin(lo)), effector memory (EM, CD45RA /CCR7 /CD27(+)/CD28(+)/ perforin(mod)) and effector (Eff, CD45RA(+)/CCR7 /CD27 / CD28 /perforin(hi)) cells. After transduction and expansion culture of peripheral blood mononuclear cells from normal donors or multiple myeloma patients, CD8(+) LeY-T cells polarized to EM-and CM-like phenotype. CD8(+) LeY-T cells differed from starting CD8(+) CM and EM T cells in that CD27, but not CD28, was downregulated. In addition, CD8(+) LeY-T cells expressed high levels of perforin, similar to starting CD8(+) Eff. CD8(+) LeY-T cells also showed hallmarks of both memory and Eff function, underwent homeostatic proliferation in response to interleukin (IL)-15, and showed interferon (IFN)-gamma production and cytotoxicity in response to Le-Y antigen on OVCAR-3 (human ovarian adenocarcinoma) cells. This study confirms CD8(+) LeY-T cells have a CM-and EM-like phenotype and heterogeneous function consistent with potential to persist in vivo after adoptive transfer. Gene Therapy (2010) 17, 1105-1116; doi:10.1038/gt.2010.59; published online 29 April 2010